Recently, researchers have been investigating the use of behavioral interventions aimed at disrupting drug- associated memories to treat addiction. One of these interventions is exposure or extinction therapy where cues associated with drugs are repeatedly presented without the drug, so that an addict learns that those cues no longer predict drug availability and they stop producing craving and relapse. Another intervention is to disrupt memory reconsolidation, where a drug cue is presented followed by an intervention designed to make the addict "forget" that the cue was associated with drug use. These methods have been used with some success to treat other psychiatric disorders. While both of these processes sound similar, there is evidence that they actually engage distinct neurobiological processes. For example, pharmacological agents that enhance extinction are also likely to enhance rather than inhibit reconsolidation. Likewise, agents that inhibit reconsolidation are also likely to inhibit extinction. In other words, the same treatment can potentially produce opposite effects on a drug memory depending on whether extinction or reconsolidation is occurring. Therefore, there is a need to find medications that can both enhance extinction and inhibit reconsolidation simultaneously, so that one does not unintentionally increase the likelihood of relapse. In this application, the candidate will conduct studies using proteomics technology to identify proteins that are differentially activated and inhibited by drug memory reconsolidation and extinction processes. The candidate has had substantial training in the behavioral pharmacology of drug addiction. The candidate has also published several papers investigating the neurobiology of both drug memory extinction and reconsolidation. However, the candidate has had little experience with the biochemical techniques required to do proteomic studies, so this proposal includes ample opportunity for structured training in proteomics, biochemistry, and molecular biology that are necessary to do these studies. The candidate's short-term career goal is to obtain training in these techniques and in the successful management of an independent laboratory. In the long-term, the candidate plans to obtain a position as a tenure-track professor at research institution to develop a broad-based, translational laboratory that will be focused on identifying and validating novel treatments for addiction. The training environment at Yale University is excellent both for the ability to complete the proposed experiments and for the training opportunities available to the candidate. Yale University has one of the leading proteomics core facilities in the nation, and experts in the latest technologies available for identifying and quantifying differences in protein phosphorylation. This is essential for the proposed experiments, as one critical component of the hypothesis is that differential activation of proteins by changes in phosphorylation state will allow the identification of novel targets for medication development to treat addiction. In addition, there are several experts within the candidate's division in the Department of Psychiatry who can provide training in biochemical and molecular biological techniques. Training at Yale University provides the advantage of several researchers in close proximity who are experts in the use of the proposed technologies in the field of addiction. The candidate will be mentored by Drs. Jane Taylor, Angus Nairn, Ralph DiLeone, and Erol Gulcicek who all provide a wealth of expertise in the research proposed. The university also provides all of the equipment and laboratory space necessary for completion of these experiments. Finally, the university provides the support, funding, and training opportunities, such as seminars and workshops, to learn new techniques and receive training in grantsmanship, lab management, mentoring, and the responsible conduct of research. The candidate proposes to use phospho-proteomics to identify novel signaling cascades that are differentially activated/inhibited by drug-cue memory reconsolidation and extinction processes. Once targets are identified, they will be validated using secondary confirmation methods including multiple reactions monitoring (MRM) and Western blotting. The targets will then be further validated in a behavioral animal model using known pharmacological agents or RNA interference via viral-mediated gene transfer. The candidate will also determine by Western blotting whether the activity of specific proteins known to be differentially activated/inhibited by reconsolidation and extinction of fear memories are also regulated by drug memories. Then, the candidate will test pharmacological or viral manipulations of these targets, which include cannabinoid receptor 1 agonists and viruses targeting inhibition of nuclear factor kappa B (NFkB) in the animal model. These experiments will provide the candidate with the opportunity to learn several new techniques and has the potential for obtaining novel information about drug memory processes that can lead to future grants as an independent scientist. The proposed experiments could also lead to the development of novel, more effective treatments for addiction that result in a long-term reduction in craving and relapse. The candidate will also spend significant time during the training period learning important skills for managing an independent laboratory, including grant writing, managing budgets, mentoring students and technicians, and training in the responsible conduct of research. PUBLIC HEALTH RELEVANCE: Addiction is a debilitating disease with significant costs to the individual and society. Successful treatments for addiction have been difficult to find, so some researchers are attempting behavioral therapies to reduce the craving caused by drug memories. The proposed studies will use proteomics to identify proteins that are activated by drug memories and will validate these proteins as targets for developing medications that could enhance the efficacy of behavioral therapies.